P8250

Peroxidase from horseradish

Type II, essentially salt-free, lyophilized powder, 150-250 units/mg solid (using pyrogallol)

• 동의어(들): Donor:hydrogen-peroxide oxidoreductase, Horseradish peroxidase
• CAS Number: 9003-99-0
• 효소 위원회 번호: 1.11.1.7 (BRENDA, IUBMB)
• EC Number: 232-668-6
• MDL number: MFCD00071339
• UNSPSC 코드: 12352204
• eCl@ss: 32160410
• NACRES: NA.54

속성

• 생물학적 소스: horseradish
• Quality Level: 300
• 유형: Type II
• 형태: essentially salt-free, lyophilized powder
• 특이 활성도: 150-250 units/mg solid (using pyrogallol)
• 분자량: ~44 kDa
• solubility: 0.1 M phosphate buffer: soluble (pH 6.0); H₂O: soluble
• 흡수비: RZ ≥1.8
• 응용 분야: diagnostic assay manufacturing
• 저장 온도: 2-8°C
• InChI: 1S/H2O3/c1-3-2/h1-2H
• InChI key: JSPLKZUTYZBBKA-UHFFFAOYSA-N

설명

일반 설명

Horseradish peroxidase is isolated from horseradish roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin group of peroxidases. HRP is a single chain polypeptide containing four disulfide bridges. It is a glycoprotein containing 18% carbohydrate. The carbohydrate composition consists of galactose, arabinose, xylose, fucose, mannose, mannosamine, and galactosamine depending upon the specific isozyme. Its molecular weight (~44 kDa) includes the polypeptide chain (33,890 Daltons), hemin plus Ca²⁺ (~700 Daltons), and carbohydrate (~9,400 Daltons). At least seven isozymes of HRP exist. The isoelectric point for horseradish peroxidase isozymes ranges from 3.0 - 9.0.

애플리케이션

Horseradish peroxidase (HRP) is isolated from horseradish roots (Amoracia rusticana). It is used in biochemistry applications such as western blots, ELISA and Immunohistochemistry. Horseradish peroxidase is used to amplify a weak signal and increase detectability of a target molecule, such as a protein. Horseradish peroxidase, product P8250, has been used to study nonoral antigens in inflamed gingiva and Ebola virus glycoprotein toxicity.

The enzyme has been used as a comparison during the peroxidase assay of extract from mature tall fescue leaf blades. It has also been used to measure H₂O₂ production.

생화학적/생리학적 작용

HRP readily combines with hydrogen peroxide (H₂O₂) and the resultant [HRP-H₂O₂] complex can oxidize a wide variety of hydrogen donors. The optimal pH is 6.0-6.5 and the enzyme is most stable in the pH range of 5.0-9.0. HRP can be conjugated to antibodies by several different methods including glutaraldehyde, periodate oxidation, through disulfide bonds, and also via amino and thiol directed cross-linkers. It is smaller and more stable than the enzyme labels β-galactosidase and alkaline phosphatase. Hence, it is the most desired label. Also, its glycosylation leads to lower non-specific binding. It is also used for the determination of glucose and peroxides in solution. Sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, and Cd²⁺, Co²⁺, Cu²⁺, Fe³⁺, Mn²⁺, Ni²⁺, Pb²⁺ ions are found to inhibit the enzyme activity.

When incubated with a substrate, horseradish peroxidase produces a coloured, fluorimetric, or luminescent derivative of the labeled molecule, allowing quantification. Horseradish peroxidase has been shown to slightly reduce the level of inhibition in a cydAB mutant.

품질

Preliminary work shows it to contain at least five isoenzymes.

단위 정의

One pyrogallol unit will form 1.0 mg purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20 °C.

분석 메모

The RZ (Reinheitszahl) is the absorbance ratio A₄₀₃/A₂₇₅ determined at 0.5-1.0 mg/ml in deionized water. It is a measure of hemin content, not enzymatic activity. Even preparations with high RZ may have low enzymatic activity.

기타 정보

View more information on peroxidase at www.sigma-aldrich.com/enzymeexplorer.

안전성 정보

• 픽토그램: GHS08
• 신호어: Danger
• 유해 및 위험 성명서: H334
• 예방조치 성명서: P261 - P284 - P501
• Hazard Classifications: Resp. Sens. 1
• Storage Class Code: 13 - Non Combustible Solids
• WGK: WGK 1
• 개인 보호 장비: Eyeshields, Gloves, type N95 (US)